Hyperactivation of FOGs formation appears to disrupt the cell wall integrity Based on the fact that cyc8Δ cells show a cell wall-defect in the mannose-media, we suspected that the cell wall integrity pathway (PKC pathway) in these cells had been compromised, and indeed PKC signaling was found to be dysregulated in cyc8Δ cells by the mannose-media 6). We further examined O-mannosylation status as well as the expression level of the cell wall sensor protein, Wsc1, which is known to be heavily O-mannosylated 7). As expected, a substantial reduction in the total amount of Wsc1 was observed in the cyc8Δ cells with the mannose-media. It was also found that the transcription level of WSC1 was not altered between WT and cyc8Δ cells after culturing them with the mannose-media, suggesting that the reduced expression level of Wsc1 in cyc8Δ cells with the mannose-media is not due to the transcriptional suppression of WSC1, but rather due to its protein instability. Collectively, these results imply that in budding yeast, the Wsc1 protein in cyc8Δ cells with the mannose-media might cause a protein instability and dysfunction in the Wsc1 protein, possibly due to the de-O-mannosylation by an EOMase 6). Taken all results together, we could successfully characterize the novel catabolic event for O-mannose glycans in budding yeast 6). EOMase has not been identified in any organisms, and therefore this study serves as a first step towards clarifying the novel catabolic pathway for O-mannose glycans. Since O-mannose glycans are conserved from yeast to human, it would be interesting to see how well this catabolic pathway is conserved in eukaryotes. Moreover, as O-mannose glycans are implicated in the genetic disorders (muscular dystrophies) in human, the identification of EOMase gene may lead to establish a valuable tool for structural analysis of human O-mannose glycans 6).Figure 2. Formation of FOGs is tightly regulated by Cyc8 6)(A) Deletion of CYC8 gene causes excessive formation of FOGs.(B) Growth of cyc8Δ cells were suppressed by the mannose-media or cell wall-perturbation reagents (calcofluor white or congo Referencesred). YPMan: mannose-media; YPGlc: control media in which mannose was replaced with glucose. 1) Hirayama H, Seino J, Kitajima T, Jigami Y & Suzuki T. Free oligosaccharides to monitor glycoprotein endoplasmic reticulum-associated degradation in Saccharomyces cerevisiae. J Biol Chem (285):12390-12404, 20102) Hirayama H & Suzuki T. Metabolism of free oligosaccharides is facilitated in the och1D mutant of Saccharomyces cerevisiae. Glycobiology (21): 1341-1348, 20113) Harada Y, Buser R, Ngwa EM, Hirayama H, Aebi M & Suzuki T. Eukaryotic oligosaccharyltransferase generates free oligosaccharides during N-glycosylation. J Biol Chem (288): 32673-32684, 20134) Hossain TJ, Hirayama H, Harada Y & Suzuki T. Lack of the evidence for the enzymatic catabolism of Man1GlcNAc2 in Saccharomyces cerevisiae. Biosci Biotechnol Biochem (80) 152-157, 20165) Hossain TJ, Harada Y, Hirayama H, Tomotake H, Seko A & Suzuki T. Structural analysis of free N-glycans in α-glucosidase mutants of Saccharomyces cerevisiae: lack of the evidence for the occurrence of catabolic α-glucosidase acting on N-glycans. PLoS One (11): e0151891, 20166) Hirayama H, et al Free glycans derived from O-mannosylated glycoproteins suggest the presence of an O-glycoprotein degradation pathway in yeast. J Biol Chem (294): 15900-15911, 2019 (Selected as “Editors’ Picks”)7) Lodder AL, Lee TK & Ballester R. Characterization of the Wsc1 protein, a putative receptor in the stress response of Saccharomyces cerevisiae. Genetics (152): 1487-1499, 199979
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